In Vivo Imaging of Myelin in the Vertebrate Central Nervous System Using Third Harmonic Generation Microscopy

Farrar, Matthew J.; Wise, Frank W.; Fetcho, Joseph R.; Schaffer, Chris B.

doi:10.1016/j.bpj.2011.01.031

Cited by 162 publications

(140 citation statements)

References 48 publications

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“…181 THG occurs even in centrosymmetric structures and can visualize interface heterogeneities as well as myelin sheaths in studies of the nervous system in animal models. 182 Since SHG and THG do not excite coupling of electronic levels, photobleaching can be suppressed, enabling observation of structures over extended periods. Squier et al 162 demonstrated THG microscopy with dynamic living specimens for the first time in 1998 with a 100 fs excitation pulse at 1.2 μm and 250 kHz repetition rate resulting in a signal at 400 nm arising from the cytoblasmic streaming and statolith movement of the chara plant rhizoids.…”

Section: Multimodal Oct and Mpmmentioning

confidence: 99%

Optical coherence tomography today: speed, contrast, and multimodality

Drexler¹,

Li²,

Kumar³

et al. 2014

J. Biomed. Opt

403

264

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Abstract. In the last 25 years, optical coherence tomography (OCT) has advanced to be one of the most innovative and most successful translational optical imaging techniques, achieving substantial economic impact as well as clinical acceptance. This is largely owing to the resolution improvements by a factor of 10 to the submicron regime and to the imaging speed increase by more than half a million times to more than 5 million A-scans per second, with the latter one accomplished by the state-of-the-art swept source laser technologies that are reviewed in this article. In addition, parallelization of OCT detection, such as line-field and full-field OCT, has shortened the acquisition time even further by establishing quasi-akinetic scanning. Besides the technical improvements, several functional and contrast-enhancing OCT applications have been investigated, among which the label-free angiography shows great potential for future studies. Finally, various multimodal imaging modalities with OCT incorporated are reviewed, in that these multimodal implementations can synergistically compensate for the fundamental limitations of OCT when it is used alone. © The Authors. Published by SPIE under a Creative Commons Attribution 3.0 Unported License. Distribution or reproduction of this work in whole or in part requires full attribution of the original publication, including its DOI.

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Section: Multimodal Oct and Mpmmentioning

confidence: 99%

Optical coherence tomography today: speed, contrast, and multimodality

Drexler¹,

Li²,

Kumar³

et al. 2014

J. Biomed. Opt

403

264

View full text Add to dashboard Cite

show abstract

“…Therefore, THG might be useful to trace myelin loss and recovery in live tissue; for example, the degeneration of myelin sheets in multiple sclerosis, which causes deficits in sensory and motor functions of the demyelinated nerves. In the brain, THG signals correspond to regions of white matter, such as the mossy fibers in the cerebellar lobes, the pons and the hindbrain, and axon bundles in the corpus callosum (Farrar et al, 2011). Dense gray matter also produces THG, with a less intense and more homogeneous signal compared to that produced by white matter (Witte et al, 2011).…”

Section: Brain and Nervous Systemmentioning

confidence: 99%

Third harmonic generation microscopy of cells and tissue organization

Weigelin

Bakker

Friedl

2016

Journal of Cell Science

167

147

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The interaction of cells within their microenvironmental niche is fundamental to cell migration, positioning, growth and differentiation in order to form and maintain complex tissue organization and function. Third harmonic generation (THG) microscopy is a label-free scatter process that is elicited by water-lipid and water-protein interfaces, including intra-and extracellular membranes, and extracellular matrix structures. In applied life sciences, THG delivers a versatile contrast modality to complement multi-parameter fluorescence, second harmonic generation and fluorescence lifetime microscopy, which allows detection of cellular and molecular cell functions in threedimensional tissue culture and small animals. In this Commentary, we review the physical and technical basis of THG, and provide considerations for optimal excitation, detection and interpretation of THG signals. We further provide an overview on how THG has versatile applications in cell and tissue research, with a particular focus on analyzing tissue morphogenesis and homeostasis, immune cell function and cancer research, as well as the emerging applicability of THG in clinical practice.

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“…Third harmonic generation (THG) microscopy is based on another nonlinear optical process of light emission, yielding distinguishable images from CARS. Though it has been demonstrated for imaging the white matter in the brain (8), so far few studies have applied THG microscopy (THGM) for elucidating the mechanism of myelin formation. Moreover, the omission of the peripheral nervous systems (PNS) in the previous studies is not trivial considering the significant departure from the CNS in terms of the myelin-forming glia and molecular subdomains: Schwann cells wrap individual internodes in the PNS, and oligodendrocytes form multiple myelin sheaths in the CNS.…”

mentioning

confidence: 99%

Label-free imaging of Schwann cell myelination by third harmonic generation microscopy

Lim¹,

Sharoukhov²,

Kassim

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

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Understanding the dynamic axon-glial cell interaction underlying myelination is hampered by the lack of suitable imaging techniques. Here we demonstrate third harmonic generation microscopy (THGM) for label-free imaging of myelinating Schwann cells in live culture and ex vivo and in vivo tissue. A 3D structure was acquired for a variety of compact and noncompact myelin domains, including juxtaparanodes, Schmidt-Lanterman incisures, and Cajal bands. Other subcellular features of Schwann cells that escape traditional optical microscopies were also visualized. We tested THGM for morphometry of compact myelin. Unlike current methods based on electron microscopy, g-ratio could be determined along an extended length of myelinated fiber in the physiological condition. The precision of THGMbased g-ratio estimation was corroborated in mouse models of hypomyelination. Finally, we demonstrated the feasibility of THGM to monitor morphological changes of myelin during postnatal development and degeneration. The outstanding capabilities of THGM may be useful for elucidation of the mechanism of myelin formation and pathogenesis.myelin | Schwann cell | multiphoton microscopy | label-free imaging | morphometry M yelin is a multiple-layered membrane sheath surrounding the axon. In myelinated nerves, the conduction of action potentials is much faster and the speed depends strongly on the structure of myelin. The structural integrity must be therefore tightly regulated for proper conduction of neuronal impulses, but the underlying axon-glial cell interaction is not well understood. Since the days of Ramón y Cajal, light microscopy has been widely used to visualize myelin morphology (1). A variety of fluorescent probes specifically binding to myelin components have allowed studies of the interaction between molecules (2-4). However, most such labeling methods are not suitable for unraveling in vivo dynamics of myelination because cell membranes are compromised during staining (especially immunohistochemistry) and/or the procedures are prohibitively timeconsuming and invasive. It is thus of great interest to develop label-free methods. Recently, spectral confocal reflectance microscopy has been demonstrated for high-resolution in vivo imaging of myelin (5). There are also techniques of nonlinear optical microscopy, which are generally known to be more advantageous for imaging deep live tissue. Coherence anti-Stokes Raman scattering (CARS) microscopy, which requires two synchronized short pulse lasers for excitation, has been used for imaging in vivo myelin and detecting pathology (6, 7). Third harmonic generation (THG) microscopy is based on another nonlinear optical process of light emission, yielding distinguishable images from CARS. Though it has been demonstrated for imaging the white matter in the brain (8), so far few studies have applied THG microscopy (THGM) for elucidating the mechanism of myelin formation. Moreover, the omission of the peripheral nervous systems (PNS) in the previous studies is not trivial considering the s...

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In Vivo Imaging of Myelin in the Vertebrate Central Nervous System Using Third Harmonic Generation Microscopy

Cited by 162 publications

References 48 publications

Optical coherence tomography today: speed, contrast, and multimodality

Optical coherence tomography today: speed, contrast, and multimodality

Third harmonic generation microscopy of cells and tissue organization

Label-free imaging of Schwann cell myelination by third harmonic generation microscopy

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